Molecular mechanism of modified Huanglian Wendan decoction in the treatment of polycystic ovary syndrome

To investigate the mechanism of modified Huanglian Wendan decoction in the intervention of polycystic ovary syndrome (PCOS) by network pharmacology and molecular docking. The ingredients and targets of modified Huanglian Wendan decoction were retrieved from the traditional Chinese medicine Systems Pharmacology database. Related targets of PCOS were screened by Comparative Toxicogenomics Database database. Cytoscape 3.7.2 (https://cytoscape.org/) was used to draw the target network diagram of “traditional Chinese medicine - ingredient - PCOS,” STRING database was used to construct the target protein interaction network. NCA tool of Cystoscape 3.7.2 was used to carried out topology analysis on PPI network, core components and key targets were obtained. Gene ontology and Kyoto encyclopedia of genes and genomes enrichment analysis were carried out for the intersection targets by David database. AutoDockTools 1.5.6 software (https://autodock.scripps.edu/) was used to conduct molecular docking verification of key components and key targets. Ninety-one ingredients of the modified Huanglian Wendan decoction and 23,075 diseases targets were obtained, 155 Intersection targets of the drug and disease were obtained by R language, Veen plot was drawn. Gene ontology enrichment analysis obtained 432 biological processes, 67 cell components, 106 molecular functions. Fifty-four Kyoto encyclopedia of genes and genomes enrichment pathways (P < .05) including tumor necrosis factor, hypoxia-induced factors-1, calcium, and drug metabolism-cytochrome P450 signaling pathway. Molecular docking showed quercetin, luteolin, kaempferol, and baicalein were stable in docking with core targets. Network pharmacology and molecular docking were used to preliminarily study the mechanism of action of modified Huanglian Wendan decoction in the treatment of PCOS, which laid foundation for future experimental research and clinical application.


Introduction
Polycystic ovary syndrome (PCOS) is the most common gynecological endocrine disease in women of reproductive age, characterized by reproductive disorders, endocrine abnormalities and metabolic disorders. [1] Its clinical manifestations are mainly irregular menstruation and infertility, the incidence of miscarriage, hirsutism, obesity and acne is as high as 5% to 10% in China, and 50% to 70% in infertile patients. [2] At present, the pathogenesis of PCOS is complex and the exact etiology is unclear, involving many aspects such as genetic, environmental and psychological factors.
PCOS belongs to traditional Chinese medicine menstruation delay, amenorrhea or hypomenorrhea. Huanglian Wendan Decoction is from "Liuyin Tiaobian of Heat Stroke," according to modern pharmacology that has significant lipid-lowering, hypoglycemic, anti-inflammatory and other pharmacological effects. [3] Tutor in the clinical flexible use of different diseases with the concept of treatment, ingenious modified, cuscutae semen, cyperi rhizoma, poria cocos, pericarpium citri reticulatae, coptidis rhizoma, angelicae sinensis radix, persicae semen, aurantii fructus immaturus, and pinellia ternata, follow the principle of compatibility, the treatment of PCOS has a good effect, but its specific mechanism of action needs further study.
Network pharmacology is an emerging bioinformatics approach in recent years to study target molecules. Through biological function and biological active compounds of traditional Chinese medicine (TCM), combined with its mechanism of action in treating diseases, to generate the complex interaction of network. The manuscript adopted the research methods of network pharmacology, and molecular docking, analysis of modified Huanglian Wendan Decoction potential targets for the treatment of PCOS and mechanism of action, for further research on modified Huanglian Wendan Decoction treatment of PCOS provides theoretical basis (Fig. 1).

Collecting targets of PCOS
Enter the keyword "polycystic ovary syndrome" into the the comparative toxicogenomics database (CTD) database [5] (http:// ctdbase.org/) to obtain the known target information related to this disease. Uniprot [6] (https://www.rcsb.org/) database was used to standardize the names of target genes.

Intersection target acquisition
The intersection of drugs component targets and diseases were obtained by R language and Venn diagram was drawn.

Ingredients-targets-TCM network
The targets of intersection between modified Huanglian Wendan decoction and PCOS were taken to establish the network diagram using Cytoscape 3.7.2, [7] the topological properties of network were analyzed. To better clarify the mechanism of the treatment of PCOS with modified Huanglian Wendan Decoction, the core network was selected based on topological parameters, degree and the betweenness centrality were key indexes to estimate the importance of nodes, as the degree and betweenness centrality increase, the nodes are more important in the network.

Target protein interaction analysis
The target at the intersection of TCM ingredients and diseases was imported into the STRING database [8] (http://string.embl. de/), the species was set as "Homo Sapiens," the lowest interaction threshold was set as "medium confidence," and the target protein interaction network was obtained.

Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis
DAVID (https://david.ncifcrf.gov/) was used to GO [9] and KEGG pathway enrichment [10] analysis on the intersection target proteins, obtained the signal pathways of modified Huanglian Wendan decoction for the intervention of PCOS.

Docking verification of core target molecules
The Cytoscape3.7.2 CNA widget was used to screen the core targets and effective active compounds according to the degree value. The chemical structure of active components in MOL2 format of TCMSP was retrieved from the database, which was re-retrieved in Pubchem database and saved in SDF format [11] (https://pubchem.ncbi.nlm.nih.gov/). Search the PDB database (https://www.rcsb.org/) for the protein and its crystal structure. After dehydration and hydrogenation, the protein was stored in PDB format with Autodoc 1.5.6 software (https:// autodock.scripps.edu/). PyMol software [12] was used to analyze the important proteins in the active ingredients and target network whose degree value was greater than the average.

Screening of common targets between compound ingredients and diseases
Data on PCOS-related targets retrieved from CTD databases were integrated, resulting in the retrieval of 23,075 targets. These targets were compared with the predicted targets, and 155 common targets were filtered as the key targets for testing the anti-PCOS activity of the ingredients (Fig. 2, Supplemental Digital Content 4, http://links.lww.com/MD/I639).

PPI network analysis
Will intersect the target were input into STRING, remove the unconnected target, the PPI network was obtained, save the TSV file. Used Cytoscape 3.7.2 for further analysis, showed that the network contained 153 nodes and 1691 edges. Network analysis was carried out through Network Viz, and 27 core targets with an average degree value >36 were identified ( Table 2). The network diagram of core targets and noncore targets was constructed ( Fig. 4A-C). Finally, different HUB computing methods were used to calculate the top 10 genes (Table 3), and it can be concluded that interleukin-6 (IL-6), vascular endothelial growth factor A (VEGFA), epidermal growth factor receptor (EGFR), mitogen-activated protein kinase 8 (MAPK8), recombinant cyclin D1 (CCND1), caspase 3 (CASP3) and FOS are common genes after HUB algorithm, and the screened active ingredients will be molecular docked.

GO functional enrichment analysis and KEGG pathway analysis
To further explore the various mechanisms of the anti-PCOS activity of modified Huanglian Wendan decoction, we performed a GO enrichment analysis, obtained 432 biological processes (BP)   selected and represented by a bar chart (Fig. 5). Among them, BP involves: response to drug, response to hypoxia, adenylate cyclase-activating adrenergic receptor signaling pathway, positive regulation of cell proliferation, synaptic transmission, cholinergic, oxidation-reduction process, positive regulation of nitric oxide biosynthetic process, positive regulation of vasoconstriction, positive regulation of extracellular regulated protein kinases1 (ERK1) and extracellular regulated protein kinases2 (ERK2) cascade. CC involves: plasma membrane, extracellular space, postsynaptic membrane, integral component of plasma membrane, synapse, caveola, axon terminus, membrane raft, mitochondrion, cytosol, cell junction, cell surface, γ-aminobutyric acid (GABA)-A receptor complex, extracellular exosome. MF involves: drug binding, enzyme binding, steroid hormone receptor activity, protein homodimerization activity, RNA polymerase II transcription factor activity, ligand-activated sequence-specific DNA binding, G-protein coupled acetylcholine receptor activity, steroid binding, extracellular ligand-gated ion channel activity, oxidoreductase activity, heme binding, serine-type endopeptidase activity, dopamine binding, dopamine neurotransmitter receptor activity. KEGG pathway enrichment analysis at P < .05 significance level was performed for the 91 modified Huanglian Wendan decoction targets. In the bubble diagram, the abscissa represents the P value, the color also reflects the enrichment degree, and the bubble size represents the number of genes. The annotation of the KEGG pathway is shown (Figs. 6A and 6B, Table 4, Supplemental Digital Content 8, http://links.lww.com/MD/ I643), mainly include: neuroactive ligand receptor interaction, tumor necrosis factor (TNF) signaling pathway, hypoxia-induced factors (HIF)-1 signaling pathway, Calcium signaling pathway, drug metabolism-cytochrome P450.

Molecular docking verification
Molecular docking is a technique that mimics the interaction between small ligand molecules and receptor protein macromolecules, and the binding energy between the 2 counterparts can be calculated to predict their affinity. A binding energy lower than 0 indicates that the 2 molecules combine spontaneously and that smaller binding energies lead to more stable conformations.  In order to further verify the prediction of the results, and elaborate the mechanism and scientific connotation of modified Huanglian Wendan decoction in the treatment of PCOS, the 4 components with the highest content of modified Huanglian Wendan decoction were selected for molecular docking verification, namely quercetin, kaempferol, luteolin, and baicalein (Table 5). IL-6, VEGFA, EGFR, MAPK8, CCND1, CASP3 and FOS were selected from RCSB protein data, direct docking was performed with grid center 0.066, 0.263,0.166, and NPTS 126 94 108 0.341. The docking results see Figure 7 and Table 6.

Discussion
Through the 3.4 and 3.5, obtained the active ingredients of modifed Huanglian Wendan decoction for treating PCOS were identified as quercetin, luteolin, kaempferol and baicalein. Modern pharmacological studies showed that quercetin could inhibit TLR/ NF-κB signaling pathway and MAPK signaling pathway, reduce the expression of testosterone (T), estradiol (E2), IL-6 and TNF-α, and increase the expression of FSH, thus alleviating the hormone, metabolism, and ovulation abnormalities caused by PCOS, it can also significantly reduce PCOS-IR and induce expression of GLUT4 and ERa genes in the uterus, therefore effectively treating PCOS and its complications: insulin resistance (IR) and infertility. [13,14] Luteolin was found to regulate the ovarian and hormone levels of rats, up-regulate Nrf2 signaling pathway, and improve antioxidant response, which is of great significance for the prevention and treatment of PCOS. [15] Studies have shown that kaempferol regulated MAPK signaling pathway to reduce inflammatory response and apoptosis, PI3K signaling pathway is mediated to promote meiosis of oocytes. [16] Baicalein regulation of InsR/IRS-1 signaling pathway, inhibits gluconeogenesis of hepatocytes to improve glucose metabolism, has strong anti-IR effect, [17] it restore the dynamic balance of reproductive hormones and normal pregnancy micro-environment through immune regulation, endocrine hormone action network, and improve the antioxidant capacity of mouse ovarian tissues by up-regulating TrxR expression. [18,19] Molecular docking, according to the results of key ingredients and targets combination conformation stability, speculated that these ingredients may be the key ingredients and targets of modified Huanglian Wendan decoction in the treatment of PCOS.
PPI results showed that IL-6, VEGFA, EGFR, MAPK8, CCND1, CASP3, and FOS showed the highest correlation among the core targets. Molecular docking proved that quercetin, luteolin, kaempferol, and baicalin had stable binding to these core targets, suggesting that network pharmacology has a high reliability for mechanism analysis of modified Huanglian Wendan decoction in the treatment of PCOS. IL-6 is a group of cytokines secreted by granulosa cells (GC), Studies have shown that IL-6 plays different roles in follicular generation, ovulation and luteal function, and is involved in the local inflammatory immune response of ovary, which is an influential factor of non-ovulation in PCOS patients. [13,20] Studies have shown that VEGFA is expressed in ovarian GC and membrane cells, as well as in follicular fluid. Abnormal VEGFA expression leads to irregular angiogenesis, which increased ovarian interstitial vascularization and perifollicular angiogenesis, involved in the development and progression of PCOS. In addition, it affects the quality of follicular mature oocytes, during pregnancy and embryo growth and development, high VEGFA concentration in follicular fluid in patients with PCOS can lead to an increase in immature follicles and a decrease in fertilized eggs. [21][22][23] EGFR is the most abundant expression in human GC and follicular fluid, has been shown to be an ovulation induction factor. When EGFR and P-EGFR are down-regulated, the induction of oocyte maturation is significantly inhibited; EGFR can also delay the meiosis of oocytes by inducing ERK1/ERK2 phosphorylation, thereby regulating ovulation time. [24][25][26] MAPK8 is a member of the protein mitokine family, which is involved in cell proliferation, cell differentiation, cell apoptosis, and other mechanisms, meanwhile, acts as an integration point for multiple biochemical signals. [27,28] Experiments have confirmed that MAPK8 is closely related to obesity and IR, expression increase in IR in obese mice, higher expression of MAPK increase the start-up process of adipocyte differentiation, promote mature adipocyte differentiation; MAPK8 known as JNK1 gene, involved in initiating the signaling pathway of JNK1/SAPK1 of cell cycle, studies have shown that IR and hyperlipidemia in PCOS patients are related to the high expression of JNK1 mRNA caused by JNK1, speculated that it may be the factor affecting the pathogenesis of PCOS. [29,30] CCND1 regulates cyclinD1 protein and promotes cytoplasmic proliferation, Cyclin D1 and its inhibitors are associated with GC proliferation at specific follicular stages, CCND1 promoted E2 secretion of PCOS ovarian GC, increased the expression level of CYP19A1, inhibited the expression level of Caspase 3 in apoptotic GC, and reduced CASP3 cleavage. [31,32] CASP3 is a regulatory factor of apoptosis signaling pathway, the early ovarian stage of PCOS patients is characterized by increased number of follicles, and the apoptosis rate of sinus follicles is related to increased expression of CASP3 in GC. Hyperandrogenemia in the late stage is closely related to excessive GC apoptosis. GC is the only cell type that can express FSH receptor, and a large number of GC apoptosis occurs. In the process of T conversion to estrogen, androgen will gradually accumulate, and CASP3 will be overexpressed. Similarly, an excess of androgens can lead to an increase in CASP3 and GC, thus leading to a sustained increase in PCOS hyperandrogenemia. [33][34][35][36][37] FOS is nuclear phosphoprotein produced by c-FOS transcription, which plays an important role in regulating cell proliferation, differentiation and apoptosis. [38] The silencing of FOS expression in GC resulted in a significant increase of CYP17 mRNA, suggesting that the FOS family regulates ovarian androgen production. [39,40] FOS can down-regulate the expression of CYP17 in cell membranes and particles, Inhibition of FOS expression in GC resulted in a significant increase in CYP17 expression and androstenedione production, even in this typical non-androstene-derived cell type [41] , The abnormal down-regulation of FOS expression in androgen-producing tissues is the potential mechanism of excessive androgen production in PCOS. FOS is the proximal promoter of FSH subunit (FSHB), regulates the expression of FSHB gene. FOS with high expression can induce FSHB (31), FOS expression is low in PCOS patients, while FSH low level is general in PCOS patients, FOS is also involved in p38MAPK/ERK1/2 signal transduction and can be induced by GnRH in gonadotropins, which proves that GnRH regulation of FSHB gene expression may be induced by FOS, [42,43] in addition, some studies have confirmed that the expression of FOS is positively correlated with the expression of inflammatory adipocytokine (IL-6, IL-8) genes, and the mRNA level of the hormone producing factor CYP19A1 is also closely correlated with FOS mRNA (39,40), therefore, it is speculated that the high expression of FOS in PCOS may have an anti-inflammatory effect and have a certain influence on steroid production in adipose tissue, inhibiting the expression of FOS may accelerate the overall development of PCOS, FOS expression was inhibited to accelerate the development of PCOS. [44][45][46] The functional enrichment results of GO biological process showed that the biological process of modified Huanglian Wendan decoction in treating PCOS mainly focused on oxidative stress, cell proliferation, Regulation of gated ion channels, and other aspects and other aspects. Hypoxia affect human health through regulating system and cell physiology of oxygen can produce reactive oxygen species, cause oxidative stress, and by changing the mammalian ovary oocytes and the physiological function of impact women's reproductive health, hypoxic conditions induced by a variety of apoptosis signaling pathways, such as autophagy and apoptosis of ovarian follicle mammal necrosis, hypoxia inducing autophagy is through 5' Amp-activated protein kinase-mammalian target of rapamyciner stress/unfolding protein response and protein kinase C-delta-C-JUNn terminal kinase 1 signaling pathway are performed in a variety of somatic cells, [47] The follicle undergoes many structural changes during growth, including changes in the vasculature cell proliferation and differentiation and formation of fluid-filled cavities, and these changes together create a hypoxic environment within the follicle so that the oocyte itself develops in a potentially hypoxic environment Survival of hypoxic tissues is controlled by HIF, which are activated in hypoxia. Understanding of the HIF signaling pathway is growing in all areas of biology, and its role in ovarian development is steadily being clarified, one of the genes upregulated by HIF is vascular endothelial growth factor, which is a major inducer of angiogenesis and is essential for ovulation and body formation. Ovulation is also intrinsically linked to HIF activity, increasing HIF expression through a surge in luteinizing hormone (LH) through ovulation. The early activation and growth initiation of dormant follicles induced by hypoxia provides evidence that these changes are associated with reduced stromal cell proliferation, suggesting that hypoxia-induced damage to the serst-cell pool may be the mechanism of accelerated follicular activation. The reversal of hypoxia can cause the decrease of SIRT1 activity in mitochondria and during hypoxia. [48][49][50] The ovarian GC of PCOS patients show abnormal proliferation, which can be reversed by reducing cell proliferation and inhibiting apoptosis with inhibitors. [51] Nitric oxide (NO) plays a role in the physiology of the reproductive system, follicular development and selection, is associated with angiogenic events, play a regulatory role in steroid production in ovarian cells by stimulating cyclooxygenase-2 activity, NO induces increased production of prostaglandin E2, which appears to be a common ovulation mechanism, NO level is negatively correlated with IR in PCOS patients, and the effects of NO and IR are mutually reinforcing, many of the effects of insulin depend on NO, which is an effector in insulin information transduction pathway. [52,53] ERK1 and ERK2 genes are closely related to IR in PCOS. The ERK2-PI3K/AKT and MAPK/ERK pathways in classic insulin-sensitive tissues target ovarian tissue, and disruption of the signaling pathway can lead to decreased insulin sensitivity and compensative hyperinsulinemia in PCOS rats. [54,55] GABA and positive-feedback GABA receptor regulatory steroids, such as isoprogesterone, stimulate food intake and weight gain. In women with PCOS, high serum isoprogeolone concentrations are associated with uncontrolled eating and disturbance of sensitivity to isoprogeolone. [56] Receptor-gated ion channels play an important role in ovarian physiology and pathology. For example, inhibition of volt-gated Ca 2+ channels can reduce extracellular Ca 2+ concentration, prevent Ca 2+ from transmembrane flow, and destroy LH stimulated steroid production in isolated ovarian cells [57] , adenylate cyclase-activating polypeptide stimulates melatonin synthesis and melatonin can increase the expression of SIRT1 to inhibit mitochondrial autophagy and protect GC from mitochondrial damage, it can also regulate dehydroepiandrosterone-induced PCOS autophagy through PI3K-Akt pathway, thereby improving ovarian dysfunction and restoring female fertility. [58][59][60] Studies have shown that GC in patients with PCOS are in an abnormal proliferation state, TNF-α activates JNK and other signal transduction pathways to promote the proliferation of follicular membrane cells, Table 4 Basic information of the top 10 pathways with the most genes.

Term
P value Genes  resulting in ovarian white membrane thickening and follicular membrane cell hyperplasia in PCOS patients, and can promote the occurrence of PCOS-IR. [61][62][63] The binding of acetylcholine (ACH) to the miAChRs receptor in GnRHergic neurons activates the phospholipase C protein, which hydrolyzes phosphatidylinositol and stimulates the formation of inositol triphosphate (IP3). The binding of IP3 to its receptor in the membrane of the rough endoplasmic reticulum moves Ca 2+ into the cytoplasm, the phosphorylated IP3 receptor decreases its response capacity when IP3 increases, induced by ACH, which results in a reduction in the release of calcium into the cytoplasm by the endoplasmic reticulum and a lower secretion of GnRH. [64] ACH and choline acetyltransferase are present in adult rats, Primates and women mature ovarian follicle in granular cells, can serve as the role of FSH partial mediator role in the growth of follicle, stimulation of GC proliferation and growth of ovarian ACH levels increase, sinus increase our secondary follicle and corpus luteum number, before the ACH to stimulate the manner and ruptured follicle and corpus luteum formation. [65,66] In conclusion, Modified Huanglian Wendan decoction exert antioxidant stress response, regulate gated ion pathway, inhibit abnormal cell proliferation and other ways to achieve IR, regulate steroid hormone disorder, reduce ovarian cell apoptosis, and finally play the role of anti-PCOS. The KEGG enrichment pathway indicated that the main pathways of midified Huanglian Wendan decoction in treating PCOS were neuroactive ligand receptor interaction, TNF signaling pathway, HIF-1 signaling pathway, Calcium signaling pathway, drug metabolism -cytochrome P450. Neuroactive ligand receptor interaction plays an important role in the reproductive process, studies have confirmed that endometrial gene expression in PCOS patients is related to neuroactive ligand receptor interaction, in addition, neuroactive ligand receptor interaction pathway is related to abnormal ovarian follicular secretion. In this pathway, IL12B and F2R genes participate in the pathophysiological mechanism of PCOS by regulating cell death and cells. Moreover, studies have confirmed that the neuroactive ligand receptor interaction pathway can regulate endocrine and emotion, and emotion regulation has obvious therapeutic effect on PCOS patients. [67,68] TNF signaling pathway is an inflammatory signaling pathway, which is involved in the pathogenesis and development of PCOS. The inflammatory factor IL-6 enriched in this pathway is involved in the local inflammatory immune response of ovary, TNF-α can inhibit FSH synthesis, and both IL-6 and TNF-α can prevent a large number of follicles from growing, Regulation of TNF signaling pathway can play an anti-PCOS role. [69][70][71][72][73] HIF-1α is a multidirectional factor that controls intracellular communication, sterogenic activity and development rate of oocytes, and affects blastocyst rate, inhibition of HIF-1α signaling pathway can not only regulate ovarian cell function, but also reduce inflammatory response and prevent oxidative stress, activation of HIF-1α signaling pathway can stimulate the expression of VEGFA gene, increase endometrial vascular hyperplasia and repair, and provide conditions for implantation of fertilized eggs. [74,75] Calcium signaling pathways involved in oxidative stress reaction, oxidative stress can promote the development of inflammation, induce IR and androgen secretion, studies have shown that calcium disorders can cause ovarian follicles block, cause reproductive and menstrual disorders, resulting in PCOS, when calcium signaling pathways activated, intracellular calcium ion concentration increases, the activation of calmodulin, prompting designed the gonadotropins release, gonadotropins indirectly regulates oocyte maturation through paracrine mechanism, resulting in ovulation, Therefore, for PCOS patients, regulating calcium ion signaling pathway can resist oxidative stress, and then play a role in resisting insulin resistance and inhibiting androgen secretio. [76][77][78][79][80] P450 aromatase is a key enzyme that catalyzes the conversion of androstenedione and T to estrone and E2, increased androgen level in serum of PCOS patients suggests that there may be defective aromatase activity, Sustained high serum levels of LH in PCOS women stimulated the production of large amounts of and rostenedione, in ovarian follicular membrane cells and stromal cells and stimulated the expression of P450 aromatase mRNA, In the study of the effect of insulin sensitizer on PCOS, it was found that metformin can directly act on the ovary, reduce the activity of P450 aromatase, reduce the synthesis of estrogen, and remove the negative feedback of estrogen on FSH in follicular fluid, increase the expression of FSH receptor and promote follicular development. [81][82][83] In summary, this study adopted network pharmacology and molecular docking methods to identify potential therapeutic targets of modified Huanglian Wendan decoction for PCOS. The relationship between modified Huanglian Wendan decoction and PCOS disease was preliminarily clarified, and the relationship between active molecules and target proteins was visualized, revealing that modified Huanglian Wendan decoction in treating PCOS is a multi-active ingredient that plays an anti-inflammatory and antioxidant stress role through multi-target and multi-pathway However, the prediction is only based on the existing database at present, which can only provide reference for the study of its mechanism of action. The subsequent research group plans to design animal experiments to conduct in-depth discussion on the mechanism of action of modified Huanglian Wendan decoction in the treatment of PCOS, so as to clarify its pharmacological mechanism more systematically and scientifically and provide more reference for its clinical use.

Conclusion
In summary, this manuscript uses network pharmacology to analyze the effective targets and pathways of adding and modified Huanglian Wendan decoction, and preliminarily explores the target action pathway and molecular mechanism of adding and modified Huanglian Wendan decoction in the treatment of PCOS. This study indicated that the mechanism of modified Huanglian Wendan decoction in treating PCOS was related to its anti-inflammatory, antioxidant stress, regulation of in vivo hormone production and resistance to insulin. The main active components are quercetin, luteolin, kaempferol, and baicalein, and the key targets are IL-6, VEGFA, EGFR, MAPK8, CCND1, CASP3 and FOS. Neuroactive Ligand receptor interaction, TNF Pathway, HIF-1 pathway, Calcium signaling pathway, Drug metabolism-Cytochrome P450 and other signal pathways are correlated, which again confirms that TCM is effective in treating diseases through multiple targets and multiple pathways. Due to many subjective and objective factors, experimental verification needs to be carried out in the next step, which will open up a new way for the treatment of PCOS. Table 6 Docking results of active ingredients and key targets (kcal/mol).